Biology Reference
In-Depth Information
mapping, and Raman-activated cell sorting have also been developed.
All these methods and developments have been recently reviewed by Li
et al.
36
and Das et al.
45
Raman scattering has been applied to microbial analysis for both food-
borne and waterborne pathogens. There is concern that culture conditions
can influence reproducibility and discrimination ability of Raman scatter-
ing, although one study has shown that even though there is some variation,
identification accuracy and taxonomic resolution are maintained.
46
Tripathi
and colleagues found that when comparing distilled and tap water, there
was no significant effect on Raman spectra and that Raman discrimination
performance was not influenced by sample aging.
47
Surface enhanced Raman spectroscopy (SERS) has been used to distin-
guish between various viruses, including adeno-, noro- and rotavirus, with
an LOD of 10
2
viruses.
48
In addition to species discrimination, viability
measurements have also been successfully demonstrated.
49
Confocal Raman
scattering has also been used for discrimination between bacteriophages
and
E. coli
. The LOD was reported as 10
9
pfu mL
−1
for bacteriophages and
10
6
cells mL
−1
, although this was considered to be due to the small sampling
volume of the confocal system with signals actually resulting from around
100 cells.
50
Bacteria quantification was attempted by Escoriza et al., who tested an
approach measuring Raman signals directly on filter materials. The chal-
lenge with this method was the choice of filter material. The substrate is
in general very important for Raman spectroscopy; given that the signal is
so weak and complex to interpret, the substrate should have a low back-
ground signal, low absorbance and high optical reflectance as well as being
nonreactive chemically. Although the membrane filters selected by these
authors were expected to give low levels of background signal, this was not
the case and high levels of bacterial coverage (10
7
cells/13 mm membrane)
were needed for detection (
Fig. 5.12
). The authors propose that future work
should concentrate on the development of new filter technologies which
can overcome these problems.
51
The authors acknowledge that Raman scat-
tering of samples smeared onto microscope slides is simpler and faster (in
terms of detection), but this obviously requires additional sample processing.
One further challenge with using membranes is that the spectra can vary
considerably between cells on membranes and those on slides (
Fig. 5.12
).
Several Raman techniques have been applied to the detection of the
protozoan,
Cryptosporidium
. The SERRS method was reported to enable
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